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311 records · Page 18
All sourcesNASA NTRSDOE OSTI
DOE OSTI2024

KBase Narrative - Lachnospiraceae sp. C1.1 genome

Narratives for The phenotype and genotype of fermentative prokaryotes This is the Narrative for Porphyromonadaceae sp. W3.11. A complementary Narrative for Lachnospiraceae sp. C1.1 is available here. This is the Narrative for Lachnospiraceae sp. C1.1. A complementary Narrative for Porphyromonadaceae sp. W3.11 is available here. Background and Isolation This Narrative and its complementary Narrative contain assembly and annotation of two bacterial isolates that were isolated by our laboratory from the rumen of a Holstein heifer. All procedures with animals have been approved by University of California Davis’s Institutional Animal Care and Use Committee. Rumen contents were collected through a rumen fistula and strained through two layers of cheesecloth into a bottle. The bottle was sealed to exclude air and maintained at 39°C. Contents were brought to the laboratory and bubbled under O2-free CO2 within 15 min. At the laboratory, serial dilutions were made with anaerobic dilution solution for Lachnospiraceae sp. C1.1 and propionibacterium diluent for Porphyromonadaceae sp. W3.11 (table S2). Aliquots (0.1 ml) of each dilution were injected into anaerobic bottle plates (1) containing 9 ml of LH medium (table S2). After incubation at 37°C for 7 days, isolated colonies were picked. Lachnospiraceae sp. C1.1 was picked from a bottle inoculated with a 104 dilution of rumen contents, and Porphyromonadaceae sp. W3.11 was picked from a bottle inoculated with a 103 dilution. After initial isolation, these organisms were purified by growing on anaerobic roll tubes (2) and picking isolated colonies. We performed de novo sequencing of Lachnospiraceae sp. C1.1 and Porphyromonadaceae sp. W3.11. Aliquots of liquid culture (9 and 1.5 ml, respectively) were collected by syringe and centrifuged (21,000g for 10 min at 4°C). Cell pellets were submitted to Molecular Research LP for DNA extraction, library preparation, and sequencing. After resuspending pellets in 180 µl of ATL buffer (Qiagen), DNA was extracted using the MagAttract HMW DNA Kit (Qiagen). DNA was eluted in 100 µl of AE buffer (Qiagen) and then cleaned using the DNEasy PowerClean Pro Cleanup Kit (Qiagen). DNA was then sheared using the Covaris g-TUBE (Covaris). Sequencing libraries were prepared using the SMRTbell Express Template Prep Kit 2.0 (Pacific Biosciences) and 1500 ng of the sheared and purified DNA. The SMRTbell libraries were size-selected (>6 Kb) using a BluePippin instrument (Sage Science) and 0.75% agarose gel. Libraries were then sequenced using the PacBio Sequel II (Pacific Biosciences) platform and a 30-hour movie time. Narrative Summary In these Narratives, we filtered low-quality reads using Trimmomatic (v0.36), assembled filtered reads with SPAdes (v3.15.3), and then checked completeness and contamination of the assembled genomes with CheckM (v1.0.18). Statistics for sequencing and assembly are in table S3. Using the assembled contigs (genomes), we called genes and annotated them. Protein-coding genes were called using Prodigal (v2.6.3) (3) locally or using KBase via RASTtk (v1.073), with identical results. Genes were annotated with KO IDs using KAAS (4). They were further annotated with pfam and TIGRFAM IDs using KBase and the Annotate Domains in a Genome app. We classified putative genes for hydrogenases using HydDB. Genes for 16S ribosomal RNA (rRNA) were called using RASTtk (v1.073) in KBase. The contigs (genomes) were analyzed to determine whether they belonged to new species. Taxonomy was assigned using GTDB-Tk (v1.7.0) in KBase. The identity of 16S rRNA genes to other organisms was found using EzBioCloud (5). Values of digital DNA-DNA hybridization (dDDH) were found with Type (Strain) Genome Server (6). These analyses suggest that Lachnospiraceae sp. C1.1 and Porphyromonadaceae sp. W3.11 represent novel species or genera. GTDB-Tk assigned Lachnospiracae sp. C1.1 to family Lachnospiraceae and genus NK4A144, which contains no type strains. It assigned Porphyromonadaceae sp. W3.11 to Porphyromonadaceae and genus Porphyromonas_A. Values of 16S rRNA identity and dDDH with respect to type strains were low (table S4). Although more phenotypic data are needed, available evidence supports assignment of genomes to new species or genera. Related publication Hackmann TJ, Zhang B. The phenotype and genotype of fermentative prokaryotes. Sci Adv. 2023 Sep 29;9(39):eadg8687. doi: 10.1126/sciadv.adg8687. Epub 2023 Sep 27. PMID: 37756392; PMCID: PMC10530074.

Hackmann, Timothy↗
DOE OSTI2021

Microbial Community Shifts Reflect Losses of Native Soil Carbon with Pyrogenic and Fresh Organic Matter Additions and Are Greatest in Low-Carbon Soils

ABSTRACT Soil organic carbon (SOC) plays an important role in regulating global climate change, carbon and nutrient cycling in soils, and soil moisture. Organic matter (OM) additions to soils can affect the rate at which SOC is mineralized by microbes, with potentially important effects on SOC stocks. Understanding how pyrogenic organic matter (PyOM) affects the cycling of native SOC (nSOC) and the soil microbes responsible for these effects is important for fire-affected ecosystems as well as for biochar-amended systems. We used an incubation trial with five different soils from National Ecological Observatory Network sites across the United States and 13 C-labeled 350°C corn stover PyOM and fresh corn stover OM to trace nSOC-derived CO 2 emissions with and without PyOM and OM amendments. We used high-throughput sequencing of rRNA genes to characterize bacterial, archaeal, and fungal communities and their responses to PyOM and OM in soils that were previously stored at −80°C. We found that the effects of amendments on nSOC-derived CO 2 reflected the unamended soil C status, where relative increases in C mineralization were greatest in low-C soils. OM additions produced much greater effects on nSOC-CO 2 emissions than PyOM additions. Furthermore, the magnitude of the microbial community composition change mirrored the magnitude of increases in nSOC-CO 2 , indicating that a specific subset of microbes was likely responsible for the observed changes in nSOC mineralization. However, PyOM responders differed across soils and did not necessarily reflect a common “charosphere.” Overall, this study suggests that soils that already have low SOC may be particularly vulnerable to short-term increases in SOC loss with OM or PyOM additions. IMPORTANCE Soil organic matter (SOM) has an important role in global climate change, carbon and nutrient cycling in soils, and soil moisture dynamics. Understanding the processes that affect SOM stocks is important for managing these functions. Recently, understanding how fire-affected organic matter (or “pyrogenic” organic matter [PyOM]) affects existing SOM stocks has become increasingly important, due to both changing fire regimes and interest in “biochar,” pyrogenic organic matter that is produced intentionally for carbon management or as an agricultural soil amendment. We found that soils with less SOM were more prone to increased losses with PyOM (and fresh organic matter) additions and that soil microbial communities changed more in soils that also had greater SOM losses with PyOM additions. This suggests that soils that already have low SOM content may be particularly vulnerable to short-term increases in SOM loss and that a subset of the soil microbial community is likely responsible for these effects.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2021

Unbalancing Symbiotic Nitrogen Fixation: Can We Make Effectiveness More Effective?

One of the most critical uses of carbon fuels is in generating nitrogen fertilizer. Atmospheric dinitrogen is too stable to be used directly by plants, so plants need other chemical forms of nitrogen. Nitrogen fertilizer production uses ~4% of the world’s natural gas. Making N fertilizers leverages methane’s energy content by enabling additional energy to be captured into food and fiber through increased photosynthesis. Manufacturing fertilizer is essential—without it there would only be enough combined nitrogen to feed ~3 billion of the world’s ~7 billion people. How to alter this addiction is not obvious, but the best chance to secure substantial and sustainable amounts of future N might be through symbiotic nitrogen fixation (SNF). SNF describes mutualistic interactions where nitrogen-fixing bacteria provide fixed nitrogen to their plant hosts, freeing them from the need for nitrogen fertilizers. SNF already occurs on a large scale; "biological nitrogen” contributes about 5 Tg N/yr to American agriculture, a quarter of the nitrogen needed. Improving SNF and replacing inorganic fertilizers are both important goals in establishing sustainable and more energy-efficient farming. SNF has been used in agriculture for millennia, largely by using legumes like beans or alfalfa in mixed cropping systems. Legume SNF occurs in root nodules, organs that develop after a growing root is infected by bacteria called rhizobia. In the plant cells of a legume nodule, specialized forms of rhizobia fix nitrogen and can produce enough ammonia to supply the growing plant. The exchange of photosynthetically derived plant carbon compounds for nitrogen reduced by the bacteria is the engine that drives the symbiosis. Important details about the organization of development and metabolism in SNF remain to be determined, such as how plant and bacterial metabolism are coordinated as nitrogen is being fixed and what governs the overall level of nitrogen fixation. Most bacterial mutations that affect SNF completely block fixation (e.g. are Fix – ), providing a limited window into the feedback interactions that must be part of the process. We have investigated an unusual mutant of Sinorhizobium meliloti, a symbiotic partner of alfalfa, that fixes dinitrogen at a normal rate (i.e., it is Fix + ) but is not effective in supporting plant growth (i.e., it is ineffective or Eff – ). We have shown that the mutant has a specific mutation in the glnD gene, which codes for a protein that regulates the bacterial response to nitrogen stress, among other key pathways. A Fix + Eff – phenotype contains an unavoidable puzzle—how is it possible for the bacteria to fix nitrogen at a normal rate and without benefiting the nitrogen starved plant host? After eliminating other possibilities, we proposed that the bacteria synthesize a nitrogen-containing compound that the plant can’t use so that acquiring this compound does not relieve plant nitrogen stress. Extracts of nodules made by glnD mutant strains have high concentrations of pyruvate canaline oxime (PCO), a derivative of the plant defensive compound canaline, a toxic analog of ornithine, an amino acid. Many legumes produce canaline and the related arginine mimic canavanine. For reasons we do not yet understand, the mutation in glnD appears to trigger significant overproduction of canaline in a defense response that leads to substantial synthesis of PCO. We expected the bacteria to be able to use PCO, but we have not been able to grow S. meliloti on PCO under several conditions. PCO thus appears to be a metabolic dead end for both the plant and bacteria. We also examined bacterial catabolic pathways that degrade arginine and canavanine and showed that mutation of these did not alter the symbiosis in an obvious way. In related experiments to examine the metabolic development of root nodules, we carried out an ambitious experiment to simultaneously measure metabolites and protein changes during nodule maturation. For these experiments we used a related symbiotic interaction between S. medicae and Medicago truncatula. M. truncatula has a simpler genome than alfalfa so we could identify plant proteins based on predictions from the DNA sequence. Nodules formed on Medicago grow linearly, with the least mature tissues at the tip. Because development of nitrogen fixation is accompanied by the production of the pigmented plant oxygen carrier protein leghemoglobin, we were able to locate and manually separate the tip, transitional, and mature nitrogen-fixing regions from each other. Advances in proteomic and metabolomic techniques allowed us to analyze these tissues from a pool of as few as 10 nodules. We observed correlated transitions of various enzymes in the plant and bacterial proteomes from those characteristic of free-living metabolism to the microaerobic metabolism of the nitrogen-fixing tissues. The metabolome could not be separated into plant and bacterial domains but was consistent with this overall transition. A subsequent paper examined proteolysis in nodule bacteria. Protein turnover in mature regions of the nodule is significant. We were able to show complexes between various proteins and important proteases, using precipitation capture techniques to isolate the proteases and sensitive proteomic analysis to reveal the associated proteins.

09 BIOMASS FUELS↗
DOE OSTI2023

The effects of protease, xylanase, and xylo-oligosaccharides on growth performance, nutrient utilization, short-chain fatty acids, and microbiota in Eimeria -challenged broiler chickens fed low-protein diet

A total of 392 Cobb 500 off-sex male broiler chicks were used in a 21-day experiment to study the effect of protease, xylanase, and xylo-oligosaccharides (XOS) on improving growth performance, nutrient utilization (ileal digestibility and total tract retention), gene expression of nutrient transporters, cecal short-chain fatty acids (SCFAs), and microbiota profile of broilers challenged with Eimeria spp. Chicks at 0-day old were allocated to 8 treatments in a 4 × 2 factorial arrangement: 1) corn-soybean meal diet with no enzyme (Con); 2) Con plus 0.2 g/kg protease alone (PRO); 3) Con plus 0.2 g/kg protease combined with 0.1 g/kg xylanase (PRO + XYL); or 4) Con plus 0.5 g/kg xylo-oligosaccharides (XOS); with or without Eimeria challenge. The 4 diets were formulated to be marginally low in crude protein (183 g/kg). Challenged groups were inoculated with a solution containing E. maxima, E. acervulina, and E. tenella oocysts on d 15. Eimeria depressed (P < 0.01) growth performance and nutrient utilization. Supplemental protease improved (P < 0.05) body weight gain and feed intake in the prechallenge phase (d 0-15) but had no effect during the infection period (d 15-21). There was no interaction between infection and feed supplementation for nutrient utilization. The supplementations of either PRO or XOS alone increased (P < 0.01) total tract retention of Ca and tended (P < 0.1) to improve total tract retention of N, P, AME, and AMEn. Eimeria decreased (P < 0.05) expressions of GLUT2, GLUT5, PepT1, ATP2B1, CaSR, Calbidin D28K, NPT2, and ZnT1 but increased (P < 0.01) expression of GLUT1. XOS supplementation increased (P < 0.05) ATP2B1 expression. Protease decreased (P < 0.05) isobutyrate concentration in unchallenged treatments but not in challenged treatments. Eimeria decreased (P < 0.01) cecal saccharolytic SCFAs acetate and propionate but increased (P < 0.01) branched-chain fatty acid isovalerate. The supplementation of PRO + XYL or XOS increased (P < 0.05) cecal butyrate or decreased cecal isobutyrate concentrations, respectively. PRO + XYL and XOS decreased cecal protein levels in unchallenged birds but not challenged ones. Eimeria challenge significantly (P < 0.05) decreased the microbial richness (Observed features) and diversity (Shannon index and phylogenetic diversity) and changed the microbial composition by reducing the abundance of certain bacteria, such as Ruminococcus torques, and increasing the abundance of others, such as Anaerostipes. In contrast, none of the additives had any significant effect on the cecal microbial composition. In conclusion, PRO or XOS supplementation individually improved nutrient utilization. All the additives decreased the cecal content of branched-chain fatty acids, consistent with decreased cecal N concentration, although the effects were more pronounced in unchallenged birds. In addition, none of the feed additives impacted the Eimeria-induced microbial perturbation.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2026

The high level trigger and express data production at STAR

To meet the demands of the Beam Energy Scan phase-II (BES-II) program, the STAR experiment at the Relativistic Heavy Ion Collider (RHIC) developed a dual real-time framework consisting of a High Level Trigger (HLT) and an Express Data Production system (xProduction). The HLT operates online within the Data Acquisition (DAQ) chain on a dedicated multi-core CPU cluster with the option to offload compute-intensive kernels to Xeon Phi coprocessors. It uses parallelized algorithms, such as the Cellular Automaton (CA) Track Finder, to perform rapid tracking, vertexing, and event filtering. This allows it to select events of interest in real time and provide immediate feedback on detector and beam conditions. In contrast, the xProduction workflow runs concurrently and independently of the DAQ loop. It applies near offline-quality calibration and reconstruction within hours of data collection. The xProduction input is the express data stream, whose content can be enriched by HLT trigger/priority selections under DAQ/HLT resource constraints, and it uses the STAR calibration/conditions framework, incorporating online calibration/QA information when available. This enables early preliminary physics analysis, including the reconstruction of rare signals, such as hyperons and hypernuclei. It also provides collaboration-wide access to analysis-ready datasets. Together, the HLT and xProduction systems form a complementary architecture: the HLT performs online event selection while the xProduction chain delivers high-quality results within a short amount of time. This integrated framework has enabled the prompt reconstruction of the $^5_Λ$ He hypernucleus with high statistical significance and the efficient processing of hundreds of millions of heavy-ion collision events. In conclusion, its demonstrated scalability and robustness establish a model for future high-luminosity experiments requiring both online event filtering and rapid access to analysis-quality data.

73 NUCLEAR PHYSICS AND RADIATION PHYSICS↗